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. Author manuscript; available in PMC: 2015 May 28.
Published in final edited form as: Science. 2014 Nov 28;346(6213):1127–1130. doi: 10.1126/science.1258973

Fig. 4. ICL unhooking.

Fig. 4

(A) The Cu-catalyzed azide-alkyne cycloaddition and the expected 1,4-isomer of the triazole ICL.

(B) Nuclease time course of the indicated ICL substrates and the unmodified control substrate (Crl-6), all in 150 mM NaCl. The uncut substrates and the products of the pre-ICL cuts (“uncut & early products”) are not resolved due to the crosslinked nature of the long oligonucleotides (fig. S8). The identity of the unhooked ICL band (“unhooked”) was confirmed by LC-MS of the band eluted from a PAGE gel of the ICL-5 reaction (table S2).

(C) Quantitation of (B), graphing the ratio of “unhooked” bands to “unhooked” plus “uncut”, corrected for the presence of two 32P-labeled strands in “uncut”. The Crl-6 curve shows total cuts, so it initially rises faster than the ICL-6 curve that does not reflect the unresolved cuts 1 and 2.

(D) Vertical lines mark incisions identified by PAGE (blue) and LC-MS (red), with line-height reflecting relative product abundance, and numbers product length (nts).